Aposematic animals use conspicuous warning signals to advertise their chemical defences to predators. Selection from predators can favour conspicuousness and large pattern elements, which enhance predator avoidance learning. In aposematic species, conspicuousness often varies among individuals. This variation can be explained if conspicuousness reflects the levels of chemical defences, if signal production or defence acquisition is costly, and if physiological trade-offs and opposing selection pressures impose constraints. To understand the link between conspicuousness and chemical defences, we must quantify the variability in warning signals and identify the chemical compounds involved. Here, we examined the warning signal variability and chemical composition of the red-necked wasp moth (Amata nigriceps). We photographed the wings of male and female moths and analysed their chemical composition using ultra-performance liquid chromatography. Females displayed more orange on their wings, a trait known to enhance protection against predators. While we ruled out the presence of pyrrolizidine alkaloids in adult moths, an untargeted metabolomics approach suggests that they sequester other compounds, such as steroidal alkaloids and alkylbenzenes, which may serve as chemical defences. Females had higher concentrations of these compounds than males but ecotoxicology assays with Daphnia showed that male and female moths exhibited similar levels of toxicity.